Tetronic, thiotetronic and tetramic acid derivatives as phospholipase A.sub.

ABSTRACT

There are disclosed compounds of the formula: ##STR1## wherein X is NR; 
     R is hydrogen or lower alkyl; 
     R 1  and R 2  are each, independently, hydrogen, C 1  -C 10  alkyl, C 3  -C 20  cycloalkyl, phenylloweralkyl, or substituted phenylloweralkyl substituted by halo, lower alkyl, lower alkoxy, halo lower alkyl, amino, monoloweralkylamino, diloweralkylamino or sulfonamido; 
     A is O, S, NR, or a chemical bond; 
     m is 0-15; 
     n is 0-20; 
     p is 0-15, 
     where m+p≦15; 
     and the pharmacologically acceptable salts thereof, which by virtue of their ability to inhibit PLA 2 , are of use as antiinflammatory agents and there is also disclosed a method for the treatment of immunoinflammatory conditions, such as allergy, anaphylaxis, asthma and inflammation in mammals, as well as in the modulation of PAF-mediated biological processes, such as embryonic implantation, thus making the compounds useful as antifertility agents.

This application is a division of U.S. application Ser. No. 08/071,627,filed Jun. 3, 1993, now U.S. Pat. No. 5,366,993, which is acontinuation-in-part of U.S. Ser. No. 07/874,928, filed Apr. 28, 1992,now abandoned.

The present invention is directed to certain tetronic acid derivativeshaving anti-inflammatory activity and to a method for using them asanti-inflammatory agents.

It is now well-established that arachidonic acid (AA) is metabolized inmammals by two distinct pathways. The metabolism of arachidonic acid bycyclooxygenase enzymes results in the production of prostaglandins andthromboxanes. The physiological activity of the prostaglandins hasalready been amply elucidated in recent years. It is now known thatprostaglandins arise from the endoperoxides PGG₂ and PGH₂ by thecyclooxygenase pathway of arachidonic acid metabolism. Theseendoperoxides are also the precursors of the thromboxanes (Tx) A₂ andB₂. TxA₂ is a vasoconstrictor which stimulates platelet aggregation. Inthe normal situation, the vasoconstrictive and platelet aggregatingproperties of the thromboxanes are balanced by another product arisingfrom the endoperoxides in the cyclooxygenase pathway, prostacyclin(PGI₂), which is a vasodilator with platelet aggregation inhibitoryactivity. In the event prostacyclin synthesis is impaired and/orplatelet activation is enhanced, then thrombosis and vasoconstriction isfavored. The role of prostanoids in haemostasis and thrombosis arereviewed by R. J. Gryglewski, CRC Crit. Rev. Biochem., 7, 291 (1980) andJ. B. Smith, Am. J. Pathol., 99, 743 (1980). Cyclooxygenase metabolitesare known to participate directly in the inflammatory response [seeHiggs et al., Annals of Clinical Research, 16, 287-299 (1984)]. This isthrough their vasodepressor activities, participation in pain and feverand augmentation of peptide mediator vascular permeability and edemaforming properties. Finally, various aspects of cell mediated immunityare influenced by cyclooxygenase products.

The other pathway of AA metabolism involves lipoxygenase enzymes andresults in the production of a number of oxidative products calledleukotrienes. The latter are designated by the LT nomenclature system,and the most significant products of the lipoxygenase metabolic pathwayare the leukotrienes B₄, C₄ and D₄. The substance denominatedslow-reacting substance of anaphylaxis (SRS-A) has been shown to consistof a mixture of leukotrienes, with LTC₄ and LTD₄ as the primary productsand having varying amounts of other leukotriene metabolites [see Bach etal., J. Immun., 215, 115-118 (1980); Biochem, Biophys. Res. Commun., 93,1121-1126 (1980)].

The significance of these leukotrienes is that a great deal of evidenceis accumulating showing that leukotrienes participate in inflammatoryreactions, exhibit chemotactic activities, stimulate lysosomal enzymerelease and act as important factors in the immediate hypersensitivityreaction. It has been shown that LTC₄ and LTD₄ are potentbronchoconstrictors of the human bronchi [see Dahlen et al., Nature,288, 484-486 (1980)], and another leukotriene, LTB₄, is a powerfulchemotactic factor for leukocytes [see A. W. Ford-Hutchinson, J. Roy.Soc, Meal.,74, 831-833 (1981)]. The activity of leukotrienes andslow-reacting substances as mediators of inflammation andhypersensitivity is extensively reviewed in Bailey and Casey, Ann.Reports Med. Chem., 17, 203-217 (1982).

Phospholipase A₂ (PLA₂) is the critical rate limiting enzyme in thearachidonic acid (AA) cascade since it is responsible for the hydrolysisof esterified AA from the C-2 position of membrane phospholipids. Thisreaction generates two products (1) free AA which is then available forsubsequent metabolism by either the cyclooxygenase or lipoxygenaseenzymes and (2) lysophospholipid. Whenalkylarachidonyl-glycerophosphatidylcholine is acted upon by the PLA₂the generation of platelet activating factor (PAF) is initiated; PAF ispro-inflammatory in its own fight [see Wedmore et al., Br. J.Pharmacol., 74, 916-917 (1981)]. In this regard it may be noted that theanti-inflammatory steroids are thought to inhibit eicosanoid synthesisby inducing the synthesis of a PLA₂ inhibitory protein denominatedmacrocortin or lipomodulin [see Flower et al., Nature, London, 278,456(1979) and Hirata et al., Proc. Natn. Acad. Sci., U.S.A., 77, 2533(1980)].

As the initial step leading to subsequent conversion of AA to thevarious eicosanoids by the cyclooxygenase and lipoxygenase pathways, thePLA₂ -mediated release of AA from membrane phospholipids is a criticalevent in attempting to deal with the various physiologicalmanifestations which are based on the activity of the eicosanoids and/orPAF. Thus, while PLA₂ has been shown to be required for plateletaggregation [Pickett et al., Biochem. J., 160, 405 (1976)], cardiaccontraction and excitation [Geisler et al., Pharm. Res. Commun., 9, 117(1977)], as well as prostaglandin synthesis [Vogt, Adv. Prostagl. Throm.Rest., 3, 89 (1978)], the inhibition of PLA₂ is indicated in thetherapeutic treatment of both PAF induced or cyclooxygenase and/orlipoxygenase pathway product-mediated physiological conditions. Thus,PLA₂ inhibitors are a rational approach to the prevention, removal oramelioration of such conditions as allergy, anaphylaxis, asthma andinflammation, as well as in the modulation of PAF-mediated biologicalprocesses, such as embryonic implantation, thus making the compoundsuseful as anti-fertility agents.

The invention provides novel compounds of the formula ##STR2## wherein Xis O, S or NR;

R is hydrogen or lower alkyl;

R¹ and R² are each, independently, hydrogen, C₁ -C₁₀ alkyl, C₃ -C₂₀cycloalkyl, phenylloweralkyl, or phenylloweralkyl substituted by halo,lower alkyl, lower alkoxy, halo lower alkyl, amino, monoloweralkylamino,diloweralkylamino or sulfonamido;

A is O, S, NR, or a chemical bond;

m is 0-15;

n is 0-20;

p is 0-15, where m+p≦15;

and the pharmacologically acceptable salts thereof.

The invention further provides a method for treating immunoinflammatoryconditions such as allergy, anaphylaxis, asthma, inflammatory boweldisease, endotoxic shock, dermatitis and psoriasis, in mammals, whichcomprises administering to a mammal so afflicted an effective amount ofa compound having the formula ##STR3## wherein X is O, S, NR,;

R is hydrogen or lower alkyl;

R¹ and R² are each, independently, hydrogen, C₁ -C₁₀ alkyl, C₃ -C₂₀cycloalkyl, phenylloweralkyl, or phenylloweralkyl substituted by halo,lower alkyl, lower alkoxy, hao lower alkyl, amino, monoloweralkylamino,diloweralkylamino or sulfonamido;

A is O, S, NR, or a chemical bond;

m is 0-15;

n is 0-20;

p is 0-15, where m+p≦15;

and the pharmacologically acceptable salts thereof.

The terms "loweralkyl" and "lower alkoxy" when used alone or incombination, refer to moieties having 1-6 carbon atoms in the carbonchain. The term "halo" refers to fluoro, chloro, bromo or iodo.

It will also be understood by those skilled in the art that thestructure: ##STR4## indicates an alkylene ring having 3+n carbon atoms.In cases where n=0, a cyclopropyl group would be indicated. It will alsobe understood that, in cases where A is a chemical bond, a continualalkylene chain is indicated in which the number of carbon atoms in thechain is equal to m plus p, i.e. equal to or less than 15.

The compounds of the invention can form pharmacologically acceptablesalts from pharmacologically acceptable organic and inorganic acids suchas hydrochloric, hydrobromic, sulfonic, sulfuric, phosphoric, nitric,maleic, fumaric, benzoic, ascorbic, pamoic, succinic, methanesulfonic,acetic, propionic, tartaric, citric, lactic, malic, mandelic, cinnamic,palmitic, itaconic and benzenesulfonic. The compounds of the inventionare capable of forming alkali metal and alkaline earth salts and saltsof pharmacologically acceptable cations derived from ammonia or a basicamine. Examples of the latter include but are not limited to cationssuch as ammonium, mono-, di-, and trimethylammonium, mono-, di-, andtriethylammonium, mono-, di- and tripropylammonium (iso and normal),ethyldimethylammonium, benzyldimethylammonium, cyclohexylammonium,benzylammonium, dibenzylammonium, piperidinium, morpholinium,pyrrolidinium, piperazinium, 1-methylpiperidinium, 4-ethylmorpholinium,1-isopropylpyrrolidinium, 1,4-dimethylpiperazinium,1-n-butylpiperidinium, 2-methylpiperidinium,1-ethyl-2-methylpiperidinium, mono-, di- and triethanolammonium, ethyldiethanolammonium, n-butylmonoethanolammonium,tris(hydroxymethyl)methylammonium, phenylmonoethanolammonium, and thelike.

The compounds within the scope of the invention by virtue of theirconfiguration, exhibit stereoisomerism. Accordingly, the compounds ofthe invention include the diastereomers, enantiomorphs, racemates andmixtures thereof.

The compounds within the scope of the invention can be prepared by avariety of synthetic routes using conventional methods. According to onepreparative scheme, a suitable cycloalkyl alcohol is oxidized to thecorresponding aldehyde, which is treated with a Wittig reagent toprovide an unsaturated ester, which in turn is reduced to the saturatedester, the latter being hydrolyzed to the free acid: ##STR5## Theintermediate free acid is then reacted with a tetronic, tetramic orthiotetronic acid to yield the desired final product: ##STR6##

In an alternative sequence, a cycloalkyl ketone is methylenated to givea epoxide which rearranges under acidic conditions to yield an aldehyde:##STR7##

The carboxylic acid starting materials (used to make the acylatedtetronic and thiotetronic acids) in which A=O or S, can be prepared bystarting with an available lipid aldehyde or ester and reducing thiscompound to the alcohol as shown below. ##STR8## To form compounds ofthis invention where A=O, the alcohol, when reacted with a suitable basesuch as sodium hydride to form the salt, is then reacted with a suitableelectrophile such as allyl bromide to give the allyl ether or withtert-butylbromoacetate to give the alkylated t-butyl ester. Thetert-buty ester can be cleaved to the acid under acidic conditions. Inthe case of the allyl ether, this compound must be oxidatively cleaved(O₃ or OsO₄ /NaIO₄) to give the aldehyde which is further oxidized tothe acid using Jones reagent or KMnO₄. See below. ##STR9## To formcompounds of this invention where A=S, the alcohol (see above), isconverted to a suitable leaving group (Br with CBr₄ /PPh₃ ; OMs withMsCl/Et₃ N, OTs with TsCl/pyridine) which is then reacted with an alkylthioglycolate salt to give the alkylated ester which can be cleaved tothe acid under saponifying conditions (base, water). ##STR10## To formcompound of this invention where A=NR, the isocyanate is prepared from asuitable lipid acid by converting to the acid chloride, reaction withTMS azide to give the acylazide. This acyl azide smoothly converts tothe isocyanate by heating. This isocyanate is then reacted with tetronicor thiotetronic acid to form the compounds of this invention. ##STR11##

The aldehyde is then subjected to olefination with a tetronic, tetramicor thiotetronic acid phosphonate, followed by hydrogenation to yield thedesired final product: ##STR12##

The starting materials in the above preparative sequences are allcommercially available or can be prepared by conventional methods astaught in the chemical literature.

The compounds of the invention, by virtue of their ability to inhibitactivity of PLA₂ enzyme, are useful in the treatment of conditionsmediated by products of the oxidation of arachidonic acid. Accordingly,the compounds are indicated in the prevention and treatment of suchconditions as allergic rhinitis, allergic bronchial asthma and othernaso-bronchial obstructive air-passageway conditions, other immediatehypersensitivity reactions, such as allergic conjunctivitis; and variousinflammatory conditions such as those present in rheumatoid arthritis,osteoarthritis, tendinitis, bursiris, psoriasis (and related skininflammation), uveitis, and the like.

When the compounds within the scope of the invention are employed in thetreatment of allergic airways disorders or in anti-inflammatory therapy,they can be formulated into oral dosage forms such as tablets, capsulesand the like. The compounds can be administered alone or by combiningthem with conventional carriers, such as magnesium carbonate, magnesiumstearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin,tragacanth, methylcellulose, sodium carboxymethylcellulose, low meltingwax, cocoa butter and the like. Diluents, flavoring agents,solubilizers, lubricants, suspending agents, binders,tablet-disintegrating agents and the like may be employed. The compoundsmay be encapsulated with or without other carriers. In all cases, theproportion of active ingredients in said compositions both solid andliquid will be at least to impart the desired activity thereto on oraladministration. The compounds may also be injected parenterally, inwhich case they are used in the form of a sterile solution containingother solutes, for example, enough saline or glucose to make thesolution isotonic. For administration by inhalation or insufflation, thecompounds may be formulated into an aqueous or partially aqueoussolution, which can then be utilized in the form of an aerosol. Thecompounds may also be used topically and for this purpose they may beformulated in the form of dusting powders, creams or lotions inpharmaceutically acceptable vehicles, which are applied to affectedportions of the skin.

The dosage requirements vary with the particular compositions employed,the route of administration, the severity of the symptoms presented andthe particular subject being treated. Treatment will generally beinitiated with small dosages less than the optimum dose of the compound.Thereafter the dosage is increased until the optimum effect under thecircumstances is reached. In general, the compounds of the invention aremost desirably administered at a concentration that will generallyafford effective results without causing any harmful or deleterious sideeffects, and can be administered either as a single unit dose, or ifdesired, the dosage may be divided into convenient subunits administeredat suitable times throughout the day.

The standard pharmacological procedures, which are described fully inthe examples given hereafter, inter alia, determine the specificity ofaction of the compounds of the invention as PLA₂ inhibitors as measuredby their ability to inhibit platelet-activating factor and LTB₄biosynthesis in human neutrophils; by their ability to inhibitarachidonic acid release mediated by human and non-human source PLA₂,and by pharmacological testing which demonstrates the ability of thecompounds of the invention to inhibit the lipoxygenase andcyclooxygenase pathways of arachidonic acid metabolism.

The following examples show the preparation and pharmacological testingof compounds within the invention.

EXAMPLE 1 3-(3-Cyclododecyl-1-oxopropyl)-4-hydroxy-2(5H)-furanone A.Cyclododecanecarboxaldehyde

To a solution of 9.92 g (50 mmol) of cyclododecane methanol in 100 ml ofmethylene chloride under a nitrogen atmosphere is added 50 mg of2,2,6,6-tetramethyl-1-piperidinyloxy, free radical (TEMPO) and 510 mg (5mmol) of sodium bromide in 5 ml of water. The solution is cooled to10°-15° C. and a solution of 100 ml of 5.25% aqueous sodium hypochloriteand 2.0 g of sodium bicarbonate is added over 1 hour. The reactionmixture is allowed to warm to room temperature and after 1 hour isdiluted with methylene chloride. The organic phase is washed withaqueous sodium thiosulfate, aqueous bicarbonate and water. The methylenechloride layer is dried over magnesium sulfate, filtered and evaporatedin vacuo to give 9.43 g of crude title compound as an oil: IR (film)1723 cm⁻¹ ; NMR (CDCl₃)) δ 1.34 (bs, 18H), 1.41 (m, 4H), 1.54 (m, 2H),1.65 (m, 2H), 2.40 (m, 1H), 9.64 (d, 1H, J=1.6 Hz) M⁺ (EI) 196.

B. 3-Cyclododecylprop-2-enoic acid, methyl ester

To a solution of 9.4 g (48 mmol) of cyclododecanecarboxaldehyde in 100ml of toluene is added 16.0 g (48 mmol) of methyltriphenylphosphoranylidene acetate. The mixture is heated to reflux andis maintained under a nitrogen atmosphere for 21 hours. The solution isremoved of solvent in vacuo and is triturated with ethyl ether to give10.8 g of white solid and a filtrate which gives an oily solid uponevaporation. The latter material is triturated with ethyl ether andfiltered to give on evaporation of the filtrate, 13.7 g of crude titlecompound: IR (film) 1722 cm⁻¹ ; NMR (CDCl₃) δ 1.36 (bs, 18H), 1.53 (m,4H), 2.35 (m, 1H), 3.73 (s, 3H), 5.78 (dd, 1H, J=1, 16 Hz), 6.88 (dd,1H, J=8, 16 Hz)

Anal. Calc'd. for C₁₆ H₂₈ O₂ : C, 76.14;H, 11.18; Found: C, 75.82;H,9.62.

C. 3-Cyclododecylpropanoic acid, methyl ester

To a solution of 12.6 g (50 mmol) of methyl 3-cyclododecylprop-2-enoatein 100 ml of methanol is added 3.6 g (150 mmol), 3 equiv) of magnesiummetal (activated by prior heating to 160° C.) and the mixture is stirredat room temperature for 2 hours (ice bath cooling is applied whennecessary to control exothermic excursions). The reaction mixture isdigested with ice cold 2N hydrochloric acid and is extracted with ethylether (3 times). The combined ethereal extracts are washed with 2Nhydrochloric acid and are dried over magnesium sulfate. Filtration andevaporation gives an oily solid. Flash chromatography on silica gelusing hexane/ethyl ether (10:1) as eluting solvent affords 7.80 g of thetitle compound: IR 1748cm⁻¹ ; NMR (CDCl₃) δ 1.33 (m, 23H), 1.54 (t, 2H,J=8 Hz), 2.31 (t, 2H, J=8 Hz), 3.66(s, 3H).

Anal. Calc'd. for C₁₆ H₃₀ O₂ : C, 75.54;H, 11.89; Found: C, 75.67;H,10.52.

D. 3-Cycledodecylpropanoic acid

To a solution of 7.6 g (30 mmol) of methyl 3-cyclododecylpropanoate in100 ml of methanol is added 2.4 g of sodium hydroxide. The mixture isheated to 55° C. and is maintained for 4 hours. The reaction mixture isdiluted with water and is extracted with hexane. The aqueous phase isacidified with 1N hydrochloric acid and is extracted (2 times) withethyl ether. The combined ethereal extracts are washed with 1Nhydrochloric acid and are dried over magnesium sulfate. Filtration andevaporation gives 7.48 g of crude title compound. Recrystallization of aportion of the product from hexane gives 1.81 g of the title compound:m.p. 72°-74° C.; IR (KBr) 1715 cm⁻¹ ; NMR (CDCl₃) δ 1.34 (m, 23H), 1.57(q, 2H, J=8 Hz), 2.36 (t, 2H, J=8 Hz). Anal. Calc'd. for C₁₅ H₂₈ O₂ : C,74.95;H, 11.74; Found: C, 74.90;H, 11.20.

E. 3-(3-Cyclododecyl-1-oxopropyl)-4-hydroxy-2(5H)-furanone

To a solution of 0.70 g (7 mmol) of tetronic acid in 20 ml of drydimethylformamide at 0° C. is added 1.1 ml (836 mg, 8.3 mmol) oftriethylamine and 0.31 g (2.5 mmol) of dimethylaminopyridine. Afterstirring for 5 minutes, 2.0 g (8.3 mmol) of 3-cyclododecylpropanoic acidis added followed by 1.6 g (8.3 mmol) of1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride. After 10minutes the ice bath is removed and the reaction mixture is allowed tostir overnight at room temperature. The reaction mixture is then pouredinto 1.0N hydrochloric acid and extracted 3 times with ethyl acetate.The organic layers are combined, washed with brine, dried over magnesuimsulfate, filtered and concentrated to give a yellow oil. The oil issubjected to flash chromatography using acidic silica gel andhexane/ethyl acetate (3:2) as the mobile phase to give an off whitesolid: m.p. 99°-102° C.; IR (KBr) 2940, 1710, 1600 and 1470 cm⁻¹ ; NMR(CDCl₃) (mixture of tautomeric forms) δ 1.34 (mc, 22H), 1.50 (bs, 1H),1.61 (m, 2H), 2.92 (m, 2H), 4.55 and 4.70 (s, 2H); MS (EI)

Anal. Calc'd. for C₁₉ H₃₀ O₄ : C, 67.87;H, 5.70; Found: C, 67.62;H,5.98.

EXAMPLE 2 3-(3-cyclododecyl-1-oxopropyl)-4-hydroxy-2(5H)-thiophenone

To a solution of 0.80 g of thiotetronic acid in 20 ml of drydimethylformamide at 0° C. is added 1.1 ml (836 mg, 8.3 mmol) oftriethylamine and 0.31 g (2.5 mmol) of dimethylaminopyridine. Afterstirring for 5 minutes, 2.0 g (8.3 mmol) of 3-cyclododecylpropanoic acidis added followed by 1.6 g (8.3 mmol) of1-(3-dimethylaminopropyl)-3-ethyl carbodiimide hydrochloride. After 10minutes the ice bath is removed and the reaction mixture is allowed tostir overnight at room temperature. The reaction mixture is then pouredinto 1N hydrochloric acid and extracted 3 times with ethyl acetate. Theorganic layers are combined, washed with brine, filtered and evaporatedto give a brown oil. The oil is subjected to flash chromatography usingacidic silica gel and hexane/ethyl acetate (9:1) as the mobile phase togive an off white solid: m.p. 60°-62° C.; IR (KBr) 2940, 1700, 1630, and1590 cm¹ : NMR (CDCl₃) δ 1.33 (bs, 22H), 1.49 (bs, 1H), 1.58 (m, 2H),2.96 (m, 2H), 3.77 and 3.98 (singlets, 2H); MS (EI) M+338.

Anal. Calc'd. for C₁₉ H₃₀ O₃ S: C, 67.87;H, 5.70; Found: C, 67.62;H,5.98.

EXAMPLE 33-(3-cyclododecyl-1-oxopropyl)-1.5-dihydro-4-hydroxy-1-methyl-1H-pyrrolin-2-oneA. 1-Oxaspiro[2.11]tetradecane

To a dry flask under nitrogen is added 3.1 g (64.6 mmol) of 50% sodiumhydride in mineral oil. The sodium hydride is washed twice with hexanes,is suspended in 80 ml of dry dimethyl sulfoxide and then 14.3 g (65.0mmol) of trimethylsulfoxonium iodide is added. The mixture is stirred atroom temperature for several hours and then 11.8 g (64.7 mmol) ofcyclododecanone is introduced. The mixture is allowed to stir overnightat room temperature and then is heated. for 1 hour at 50° C. The mixtureis cooled to room temperature, is dissolved in 500 ml of cold water andis extracted seven times with ethyl ether. The combined organic layersare washed once with water, dried over magnesium sulfate and subjectedto evaporation in vacuo. The residue is chromatographed on silica gelusing 20% ethyl acetate/hexanes as the mobile phase to give 10 g (78%)of the title compound as a colorless oil: NMR (CDCl₃) δ 1.2-1.8 (m, 18H), 2.05 (s, 1H), 2.4 (t, 2H, J=5.0 Hz), 2.45 (d, 2H), 2.58 (s, 2H ).

B. Cyclododecanecarboxaldehyde

To 5.0 g (25.5 mmol) of 1-oxaspiro[2.11]tetradecane in 50 ml of dryether at 0° C. is slowly added 1.8 ml (2.1 g, 14.6 mmol) of borontrifluoride etherate. The ice bath is removed and after 15 minutes atroom temperature the solution is partitioned between ether and saturatedsodium bicarbonate. The organic phase is evaporated to give a clear oil.Flash chromatography over silica gel using a hexane to hexane:ethylacetate (9:1) gradient give 1.6 g (34%) of a clear oil: NMR (CDCl₃) δ1.28-1.8 (m, 18H), 2.03 (s, 1H), 2.42 (m, 1H), 9.62 (s, 1H)

C. Sarcosine, N-(4-bromo-1,3-dioxo)butane, ethyl ester

To 38 ml (40.8 g, 486 mmol) of diketene in 200 ml of dry methylenechloride at -78° C. is slowly added 24.0 ml (75 g, 468 mmol) of bromine.After stirring at -78° C. for 1 hour, the mixture is subjected to theaddition of 90 ml (65.3 g, 647 mmol) of triethylamine followed by 50 g(326 mmol) of sarcosine ethyl ester hydrochloride. After stirring at-78° C. for 2 hours no starting materials are evident by thin layerchromatography. The mixture is taken up in water and the methylenechloride phase is removed. The organic layer is dried over magnesiumsulfate and evaporated to give a black residue which is subjected toflash chromatography on silica gel using hexanes/ethyl acetate (3:2) togive 23.1 g of the title compound as a crude red oil which is usedwithout further purification.

D. [2-(1,5-Dihydro-4-hydroxy-1-methyl-2-oxo-1H-pyrrol-3-yl)-2-oxoethyl]phosphonic acid, diethyl ester

To 28.0 g (250 mmol) of potassium t-butoxide in 40 ml ofdimethylformamide at 0° C. is slowly added 29 ml (31 g, 225 mmol) ofdiethyl phosphite. The mixture is stirred for 45 minutes and 20.0 g(75.2 mmol) of sarcosine, N-(4-bromo-1,3-dioxo)butane ethyl ester in 20ml of dry tetrahydrofuran is added. After the addition is complete, themixture is allowed to stir at room temperature overnight. Thin-layerchromatography using phosphoric acid-treated plates indicates theconsumption of starting material. The mixture is acidified with 1Nhydrochloric acid and is extracted three times with dichloromethane. Thecombined organic layers are taken up in 1N sodium hydroxide, washed withdichloromethane, reacidified and then washed three times withdichloromethane. The combined organic layers are dried over magnesiumsulfate and evaporated to give 19.17 g (88%) of a dark oil: NMR (CDCl₃)δ1.35 (t, 4H, J=7 Hz), 2.9 (s, 2H), 2.97 (s, 2H), 3.05 (s, 2H), 3.6 (d,1H, J=25 Hz), 3.8 (s, 1H), 4.2 (m, 4H), 8.0 (s, 1H)

E.3-(3-Cyclododecyl1-ox0-2-propenyl)-1,5-dihydro-4-hydroxy-1-methyl-1H-pyrrol-2-one

To a solution of 1.3 g (4.5 mmol) of diethyl[2-(1,5-dihydro-4-hydroxy-1-methyl-2-oxo-1H-pyrrol-3-yl)-2-oxoethyl]phosphonatein dry tetrahydrofuran at 0° C. is slowly added 1.0 g (8.9 mmol) ofpotassium t-butoxide in 20 ml of dry tetrahydrofuran. After 0.5 hour at0° C., 0.8 g (4.1 mmol) of cyclododecanecarboxaldehyde is added and thereaction mixture is allowed to stir overnight at room temperature. Themixture is evaporated under reduced pressure and the residue isdissolved in 1N sodium hydroxide, washed once with dichloromethane, andacidified with concentrated hydrochloric acid. The acidified aqueousphase is extracted four times with dichloromethane and the combinedorganic phases are dried over magnesium sulfate and evaporated in vacuo.The residue is chromatographed using acidic silica gel and hexane:ethylacetate (3:1) to give 0.56 g (35%) of an off white solid: NMR (CDCl₃) δ2.2-2.7 (bs, 22H), 2.45 (m, 2H), 3.0 (s, 3H), 3.81 (s, 2H), 7.1 (s, 2H).

F.3-(3-Cyclododecyl-1-oxopropyl)-1,5-dihydro-4-hydroxy-1-methyl-1H-pyrrolin-2-one

To 0.56 g (1.7 mmol) of3-(3-cyclododecyl-1-oxo-2-propenyl)-1,5-dihydro-4-hydroxy-1-methyl-1H-pyrrol-2-onein 20 ml of ethyl acetate is added a catalytic amount of 10% palladiumon carbon. The mixture is allowed to stir at room temperature for 5hours after which time no starting material is evident by thin layerchromatography. The mixture is passed through a Celite pad and isevaporated in vacuo. The residue is chromatographed over acid-treatedsilica gel using hexane/ethyl acetate (2:1) to give 0.45 g of a whitesolid: m.p. 66°-68° C.; IR (KBr) 2820, 2660, 1720, 1650, 1600 cm⁻¹ ; NMR(CDCl₃) δ 1.28 (bs, 22H), 1.44 (bs, 1H), 1.51 (m, 2H), 2.78 and 2.88(triplets, 2H, J=8 Hz), 2.95 and 2.97 (singlets, 3H, 1:5), 3.66 and 3.79(s, 2H, 7:1); MS M+335.

Anal. Calc'd. for C₂₀ H₃₃ NO₄ : C, 71.60;H, 9.91, N, 4.17; Found: C,71.23;H, 9.64; N, 4.14.

EXAMPLE 4 3-(3-cyclododecyl-1-oxopropyl)-4-hydroxy-2(5H-thiophenonecompound with 2-amino-2-(hydroxymethyl)-1,3-propanediol (1:1)

To 2.5 g (74 mmol) of3-(3-cyclododecyl-1-oxopropyl)-4-hydroxy-2(5H)thiophenone in 20 mL ofethanol at 70° C. is added 0.85 g (7.0 mmol) oftrishydroxymethylaminomethane dissolved in 5 mL of water. The reactionmixture is stirred at 70° C. for 0.5 hours and then the solvents areevaporated. The residue is recrystallized from tetrahydrofuran/isopropylether (2:1) to give 1.3 g of a whim solid: m.p. 164°-166° C.; IR (KBr)2860-3460, 1640, 1580 cm⁻¹ ; NMR (DMSO-d₆) δ1.2-1.4 (m, 10H), 2.55 (t,2H, J=7 Hz), 3.25 (s, 2H), 3.3 (s, 6H), 3.47 (d, 6H, J=5 Hz), 5.15 (t,3H, J=5 Hz), 7.6 ppm (bs, 3H); MS ((+)FAB): [M-Base+2H]339.

Anal. Calc'd. for C₂₃ H₄₁ NO₆ S: C, 60.1;H, 8.99; N, 3.05; Found: C,59.82;H, 8.74; N, 3.77.

EXAMPLE 5 3-(4-cyclododecyl-1-oxobutyl)-4-hydroxy-2(5H)-thiophenone A.1-(3-phenyl-1-propenyl)cyclododecane

To 11.4 g (25.5 mmol) of phenylethyl triphenylphosphonium bromide in 50mL of dry ether at 0° C. is added 10.2 mL (25.5 mmol) of 2.5Mbutyllithium. The suspension is stirred at 0° C. for 1 hour followed bythe addition of 5.0 g (25.5 mmol) of cyclododecane carboxaldehyde. Themixture is stirred at room temperature for 3 hours after which time nostarting material is evident by TLC. The reaction mixture is quenchedwith 1N HCl. The layers are separated and the aqueous layer is washedtwice with ethyl ether. The combined organic layers are dried over aMgSO₄ and the ether is evaporated. The residue is flash chromatographedon silica gel using hexanes to give 6.4 g of a colorless oil: NMR(CDCl₃) δ 1.0-1.6 (m, 10H), 2.6 (m, 1H), 3.4 (d, 2H, J=8 Hz, ), 5.3 (t,1H, J=7 Hz), 5.45 (q, 1H, J=7 Hz), 7.2 ppm (m, 5H).

B. 1-(3-phenylpropyl)cyclododecane

To 6.4 g (22.5 mmoles) of 1-(3 phenyl-1-propenyl)cyclododecane in 50 mLof ethanol is added 0.48 g of 10% palladium on carbon. The reactionmixture is subjected to hydrogen (50 psi) atmosphere using a Paarhydrogenerator and shaken overnight after which time no startingmaterial is evident by TLC. The reaction mixture is filtered throughCelite which is washed with additional ethanol. The filtrate isevaporated and the crude residue (5.8 g) is used directly for the nextstep: NMR (CDCl₃) δ 1.2-1.4 (m, 10H), 1.6 (m, 2H), 2.55 (t, 2H, J=8 Hz),7.2 (m, 5H).

C. 4-cyclododecylbutanoic acid

To a flask containing 90 mL of water, 60 mL of acetonitrile and 60 mL ofcarbon tetrachloride is added 4.8 g (16.8 mmol) of1-(3-phenylpropyl)cyclododecane, 50.0 g (0.25 mol) of sodium periodateand then 70 mg (0.3 mmol) of ruthenium trichloride hydrate. The reactionmixture is allowed to stir at room temperature overnight. The mixture isthen taken up in dichloromethane and 1N HCl and the layers separated.The aqueous phase is washed 3 times with dichloromethane and thecombined organic layers are dried over MgSO₄ and the solventsevaporated. The residue is taken up in ether, filtered through Celiteand the filtrate evaporated. The crude product is flash chromatographedusing acidic silica gel and 30% ethyl acetate/hexanes to give 3.25 g ofa white solid: NMR (CDCl₃) δ 1.2-1.5 (m, 10H), 1.65 (m, 2tt), 2.35 (t,2H, J=8 Hz).

D. 3-(4-cyclododecyl-1-oxobutyl)-4-hydroxy-2-(5H)thiophenone

To a dry flask under N2 atmosphere at 0° C. is added 1.31 g (11.3 mmol)of thiotetronic acid in 40 mL of dry dichloromethane followed by 1.42 mL(10.2 mmol) of triethylamine and 0.38 g (3.1 mmol) of4-dimethylaminopyridine. After stirring at 0° C. for 5 minutes, 2.6 g(10.2 mmol) of cyclododecylbutanoic acid is added followed by 4.8 g(11.3 mmol) of 1-cyclohexyl-3-(2-morpholinoethyl) carbodiimide methop-toluenesulfonate. After stirring for 10 minutes, the reaction mixtureis allowed to warm to room temperature and is stirred overnight. Thereaction mixture is quenched with 1N HCl. The layers are separated andthe aqueous phase is washed once with dichloromethane. The combinedorganic layers are washed twice with aqueous sodium bicarbonate andtwice with aqueous 2N HCl. The organic phase is then dried over MgSO₄and the dichloromethane evaporated. The residue is flash chromatographedon acidic silica gel using 8% ether/hexanes, dissolved in ether, washedwith 1N HCl, dried and evaporated to give 1.2 g of a white solid: m.p.62°-69° C.; IR; NMR (CDCl₃) δ 1.2-1.4 (m, 10H), 1.5 (s, 4H), 1.65 (m,2H), 2.9 (t, 2H, J=7 Hz), 3.96 and 3.75 (s, 2H); MS

Anal. Calc'd. for C₂₀ H₃₂ O₃ S: C, 68.53;H, 8.63; Found: C, 67.72;H,8.84.

EXAMPLE 6 3-(4-Cyclododecyl-1-oxobutyl)-4-hydroxy-2(SH)-thiophenonecompound with 2-amino-2-(hydroxymethyl)-1,3-propanediol (1:1)

To 0.8 g (2.3 mmol) of3-(4-cyclododecyl-1-oxobutyl)-4-hydroxy-2-(5H)thiophenone in 20 mL ofethanol at 70° C. is added 0.26 g (2.2 mmol) of trishydroxymethylaminomethane dissolved in 5 mL of water. The reaction mixture is stirredat 70° C. for 0.5 hours and then the solvents are evaporated. Theresidue is recrystallized from tetrahydrofuran/ether (2:1 ) to give 0.66g of an off white solid: m.p. 141°-144° C.; IR (KBr) 3420, 2920, 2830,1580 cm⁻¹ ; NMR (DMSO-d₆) a 1.1-1.4 (m, 10H), 2.55 (t, J=7 Hz, 2H), 3.26(s, 1H), 3.31 (s, 4H), 3.47 (d, J=5, 3 Hz), 5.15 (t, J=5 Hz, 2H), 7.6(bs, 2H); MS (DCI) [M+H]⁺ 353.

Anal. Calc'd. for C₄ H₄₃ NO₆ S: C 60.86;H 9.15; N, 2.96; Found: C,60.91;H 9.16; N, 2.88.

EXAMPLE 7A Cyclododecylene acetic acid

A solution of 12 mL, of dicyclohexylamine (60.4 mmol) in 100 mL of dryTHF was subjected to the addition of 24.2 mL of 2.5 M in butyl lithium(60.4 mmol) at -78° C. After 15 minutes, 11.1 mL of ethyl trimethylsilylacetate (60.4 mmol) was added dropwise and after another 15 minutes, 5.5g of cyclododecanone (30.2 mmol) in dry THF was added dropwise. Afterstirring at 78° C. for 1 h, the reaction mixture was allowed to warm toroom temperature. 1.2 g of sodium bisulfate monohydrate was added andthe reaction mixture was stirred for another 15 minutes. It was quenchedwith saturated NH₄ Cl followed by 1N HCl. The phases were separated, theaqueous layer washed with ether. The combined organic layers were washedwith brine, dried over MgSO₄ and evaporated to give 6.0 g of a tan oil(79%).

EXAMPLE 7B Ethyl cyclododecyl acetate

To 6.0 g of ethyl cyclodecylane acetate (23.8 mmol) in 150 mL ofmethanol was added 3.0 g (119 mmol) of magnesium turnings. The reactionmixture was stirred at room temperature overnight, after which time, allthe magnesium had been consumed. The reaction was quenched with 3N HCland extracted with ether. The combined ether layers were dried overMgSO₄ and evaporated to give 5.5 g of a tan oil (92%).

EXAMPLE 7C Cyclododecyl acetic acid

To a solution of 1.73 g of sodium hydroxide (43.3 mmol) in water wasadded 5.5 g of ethyl cyclododecyl acetate in methanol. Water and THFwere added until the solution was homogeneous. The reaction mixture washeated to 50° C. for several hours, the solvents stripped and theresidue was partitioned between ether and 0.5N sodium hydroxide. Theaqueous phase was acidified and extracted 3× with ether. The etherphases were combined, washed with brine, dried over MgSO₄ and evaporatedto give 2.8 g of a white solid.

EXAMPLE 7D 3-(2-cyclododecyl-1-oxoethyl)-4-hydroxy-2(5H)-furanone

To a dry flask under N₂ atmosphere at 0° C. is added 1.46 g (14.6 mmol)of tetronic acid in 40 mL of dry dichloromethane followed by 1.85 mL(13.3 mmol) of triethylamine and 0.49 g (4.0 mmol) of4-dimethylaminopyridine. After stirring at 0° C. for 5 minutes, 3.0 g(13.3 mmoles) of cyclododecyl acetic acid is added followed by 6.12 g(14.6 mmol) of 1-cyclohexyl-3-(2-morpholinoethyl) carbodiimide methop-toluene sulfonate. After stirring for 10 minutes, the reaction mixtureis allowed to warm to room temperature and stirred overnight. Thereaction mixture is quenched wih 1N HCl. The layers are separated andthe aqueous phase is washed once with dichloromethane. The combinedorganic layers are washed twice with aqueous sodium bicarbonate andtwice with aqueous 2N HCl. The organic phase is then dried over MgSO₄and the dichloromethane evaporated. The residue is flash chromatographedon acidic silica gel using 90% ether/hexanes, taken up in ether, washedonce with 1N HCl and recrystallized from cyclohexane to give 2.0 g of alight yellow solid: m.p. 133°14 135° C.; IR (KBr) 2920 (C--H), 1770,1650, 1600 cm⁻¹ (C=O); NMR (CDCl₃) δ 1.2-1.5 (m, 10H), 2.1 (m, 1H), 2.84(d, 2H, J=7 Hz), 4.6 (bs, 2H), MS (EI) M⁺ 308.

Anal. Calc'd. for C₁₈ H₂₈ O₄ : C, 70.10;H, 9.15; Found: C, 70.38;H,9.21.

EXAMPLE 8 3-(2-cyclododecyl-1-oxoethyl )-4-hydroxy-2 (5H )-thiophenone

To a dry flask under N2 atmosphere at 0° C. is added. 1.7 g (14.6 mmol)of thiotetronic acid in 40 mL of dry dichloromethane, followed by 1.85mL (13.3 mmol) of triethylamine and 0.49 g (40 mmol) of4-dimethylaminopyridine. After stirring at 0° C. for 5 minutes, 3.0 g(13.3 mmol) of cyclododecylacetic acid is added followed by 6.12 g (14.6mmol) of 1-cyclohexyl-3-(2-morpholinoethyl) carbodiimide methop-toluenesulfonate. After stirring for 10 minutes, the reaction mixtureis allowed to warm to room temperature and stirred overnight. Thereaction mixture is quenched with 1N HCl. The layers are separated andthe aqueous layer is washed once with dichloromethane. The combinedorganic layers are washed twice with aqueous sodium bicarbonate and thentwice with aqueous 2N HCl. The organic phase is then dried over MgSO₄and the dichloromethane evaporated. The residue is flash chromatographedon acidic silica gel using 10% dichloromethane/hexanes, taken up inether, washed once with 1N HCl and recrystallized from hexane to give1.2 g of a white solid: m.p. 85°- 87° C.; MS (EI) M⁺ 324; IR (CHCl₃)2930, 2860 (C-H), 1685, 1620, 1565 cm⁻¹ ; NMR (CDCl₃) δ 1.2-1.5 (m,10H), 2.1 (m, 1H), 2.87 (d, 2H, J=7 Hz), 3.96 and 3.75 (s, 2H).

Anal. Calc'd. for C₁₈ H₂₈ O₃ S: C, 66.63;H, 8.70; Found: C, 67.46;H,8.81.

EXAMPLE 9 3-(3-cyclohexyl-1-oxonpropyl)-4-hydroxy-2(5H)-furanone

To a solution of 2.0 g of tetronic acid in 40 ml of dryN,N-dimethylformamide was added (at 0° C.) 3.1 ml of triethylamine and0.81 g of dimethylaminopyridine. After stirring for 5 minutes, 3.4 ml of3-cyclohexylpropanoic acid was added, followed by 4.6 g of1-(3-dimethylaminopropyl)-3-ethyl carbodiimide hydrochloride. After 10minutes the ice bath is removed and the mixture is allowed to stirovernight. The reaction mixture is quenched with 1N hydrochloric acidand the solution is washed 3 times with hexanes. The combined organiclayers are combined, washed with brine, dried over magnesium sulfate andevaporated to give a yellow solid, which is recrystallized from hexanesto give light yellow crystals: m.p. 75°14 78° C.; IR (KBr) 2910, 2820,1750, 1640, 1600 cm⁻¹ ; NMR (CDCl₃) δ Ε 0.9-1.8 (1 1H, m), 2.9 (t, 2H,J=6 Hz), 4.7 and 4.5 (s, 2H (tautomers)); MS EI M⁺ 238.

Anal. Calc'd. for Cl₃ H₁₈ O₄ : C, 65.53;H, 7.61; Found: C, 65.23;H,7.74.

EXAMPLE 10 3-(3-cyclohexyl-1-oxopropyl)-4-hydroxy-2(5H )-thiophenone

In the same manner as that described for Example 1 is prepared the titlecompound with the exception that thiotetronic acid is used instead oftetronic acid. Crystallization from hexanes gives yellow crystals: m.p.:60°-62° C.; NMR (CDCl₃) δ 0.9-1.7 (m, 11H), 2.95 (t, 2H, J=8 Hz), 4.0(s, 2H); IR 2920, 2840, 1680, 1560 cm⁻¹ ; MS (El) M⁺ 254.

Anal. Calc'd. for Cl₃ H₁₈ SO₃ : C, 61.39;H, 7.30; Found: C, 60.98;H,7.30.

EXAMPLE 11 3-(4-cyclohexyl-1-oxobutyl)-4-hydroxy-2(SH)-furanone

To a solution of 1.0 g (10.0 mmol) of tetronic acid in 30 mL of dryN,N-dimethylformamide at 0° C. is added 1.5 mL (11 mmol) oftriethylamine followed by 0.37 g (30 mmol) of 4 dimethylaminopyridine.After stirring at 0° C. for 5 minutes, 2.0 g (11.7 mmol) of 4-cyclohexylbutyric acid is added, followed by 2.25 g (11.7 mmol) of1-(3-dimethylaminopropyl)-3-ethyl carbodiimide. After 10 minutes, theice bath is removed and the reaction mixture is allowed to stirovernight. The reaction mixture is quenched with 1N HCl and the solutionis washed three times with hexanes. The combined organic layers arewashed with water, dried over magnesium sulfate and evaporated. Theresidue is flash chromatographed on silica gel using 5% methanol/ethylacetate with 2 mL of acetic acid/liter to give an orange solid: m.p.38°-42° C.; IR (film) 2920, 2840, 1770, 1690, 1660, 1600 cm⁻¹ ; NMR(CDCl₃) δ 0.9 (m, 2H), 1.2 (m, 6H), 1.7 (m, 6H), 2.9 (q, J=7 Hz, 2H),4.6 and 4.7 (s, 2H), 12.1 (bs, 1H); MS (EI): M+252.

Anal. Calc'd. for C₁₄ H₂₀ O₄ : C, 66.64;H, 7.99; Found: C, 66.38;H,7.90.

EXAMPLE 12 3-(4-cyclohexyl-1-oxobutyl)-4-hydroxy-2 (5H )-thiophenone

The title compound is prepared in the same manner as that described forExample 11 with the exception that thiotetronic acid is used instead oftetronic acid. A tan solid is obtained: m.p. 45°-50° C.; IR (KBr): 2970,2840, 1660, 1590 cm⁻¹ ; NMR (CDCl₃) δ0.9 (m, 2H) 1.2 (m, 6H) 1.7 (m,6H), 2.9 (t, J=7 Hz, 2H), 3.8 and 4.0 (s, 2H), 16.1 (bs, 1H); MS (EI) M⁺268.

Anal. Calc'd. for C₁₄ H₂₀ O₃ S: C, 62.66;H, 7.51; Found: C, 62.67;H,7.53.

EXAMPLE 13A Cyclododecyl 3 allyl ether

To 2.6 g (54.3 mmol) of 50% sodium hydride that had been washed twicewith hexanes was added 15 mL of N,N-dimethyl formamide followed by 10.0g (5.4 mmol) of cyclododecanol. The reaction mixture was stirred at roomtemperature for several hours and then at 65° C. until the evolution ofgas had ceased. 9.4 mL (108.5 mmol) of allyl bromide was then added andthe reaction mixture was allowed to stir overnight at 65° C. Thereaction was quenched with water, extracted 3 times with ether, thecombined ether layers washed with water, dried over MgSO₄ andevaporated. The residue was flash chromatographed on silica gel using 2%Et₂ O/Hexanes to give 5.34 g of an oil (44% ).

EXAMPLE 13B 3(cyclododecyloxy)1.2 dihydroxy propane

To a mixture of cyclododecyl 3 allyl ether (5.0 g, 22.3 mmol) and 88%formic aced (26.3 mL) was added 3.12 g (24.5 mmol) of 30% hydrogenperoxide. The reaction mixture was heated to 40° C. and allowed to stirovernight. The excess formic acid and water was removed by rotaryevaporation and the residue was refluxed for 1 h in 3N ethanolicpotassium hydroxide. The ethanol was removed and the residue partitionedbetween ether and 1N HCl. The aqueous layer was washed with ether andthe combined ether layers were dried over MgSO₄ and evaporated. Theresidue was flash chromatographed on silica gel using 70% ethylacetate/hexanes to give 3.58 g of an oil (62%). NMR (CDCl₃): 3.4-3.8ppm(m, 6H); 3.2 (bs, 2H); 1.4 (m, >10H).

EXAMPLE 13C Cyclododecyloxy acetic acid

To 1.9 g (7.4 mmol) of 3(cyclododecyloxy)1,2 dihydroxy propane in 20 mLof benzene was added 3.3 g (7.4 mmol) of lead tetraacetate and 2.0 g(14.8 mmol) of potassium carbonate. After stirring for several hours atroom temperature, the reaction mixture was partitioned between ether and1N HCl. The aqueous layer was washed with ether and the combined etherlayers were dried over MgSO₄ and evaporated.

The crude residue was subjected to oxidation by Jones reagent in acetoneat 0° C. The reaction n-fixture was again partitioned between ether and1N HCl. the aqueous layer was washed with ether and the combined etherlayers dried over MgSO₄ and evaporated. The residue was flashchromatographed on acidic silica gel using 10% EtOAc/Hexanes to give 0.7g of a white solid (40%). IR (film): 2950, 2880, 1760, 1480 cm⁻¹ NMR(CDCl₃): 3.7 (s, 3H); 3.15 (s, 3H); 2.4 (t, 2H, J=8 Hz); 1.55 (q, 2H,J=6 Hz); 1.3 (m, >10H); MS(EI): M⁺ 283;

    ______________________________________                                                         Theory                                                                              Found                                                  ______________________________________                                        Analysis: % C          72.03   71.13                                                    % H          11.73   11.48                                                    % N          4.94    4.17                                           ______________________________________                                    

EXAMPLE 13D 3-(2-cyclododecyloxy-1-oxoethyl)-4-hydroxy-2(5H)-thiophenone

To a solution of 0.37 g (3.2 mmol) of thiotetronic acid in 20 mL of drydichloromethane at 0° C. is added 0.4 mL (2.9 mmol) of triethylaminefollowed by 0.11 g (0.9 mmol) of 4-dimethylaminopyridine. After stirringat 0° C. for 5 minutes, 0.7 g (2.9 mmol) of cyclododecyloxyacetic acidis added, followed by 1.35 g (3.2 mmol) of1-cyclohexyl-3-(2-morpholinoethyl) carbodiimidemetho-p-toluenesulfonate. After 10 minutes the ice bath is removed andthe reaction mixture is allowed to stir overnight. The reaction mixtureis quenched with 1N HCl. The layers are separated and the organic layeris washed twice with aqueous sodium bicarbonate and then twice with 2NHCl. The organic phase is then dried over MgSO₄ and the dichloromethaneevaporated. The residue is flash chromatographed on acidic silica gelusing 50% ether/hexanes and then recrystallized from hexanes to give ayellow solid: m.p. 93°-95° C.; IR (KBr) 2920, 2840, 1690, 1660, 1580cm⁻¹ ; NMR (DMSO-d₆) δ 1.2-1.6 (m, 10H), 3.4 (m, 1H), 3.95 and 4.4 (s,2H), 4.6 (bs, 2H); MS (EI) M+340.

Anal. Calc'd. for C₁₈ H₂₂ O₄ S: C, 63.50;H, 8.29; Found: C, 63.30;H,8.41.

EXAMPLE 14 2-Cyclododecylethyl isocyanate

To 5.0 g of 3-cyclododecylpropionic acid (20.8 mmol) is 40 mL of toluenewas added 7.6 mL (100 mmol) of thionyl chloride. The solution wasrefluxed overnight, stripped of solvent, taken up in toluene, and againstripped. The residue was taken up in toluene, subjected to the additionof 8.4 mL of trimethylsilyl azide (60 mmol), and heated to 100° C. untilno further evolution of gas was observed. The reaction mixture wasstripped of solvent and the crude product was used directly for the nextreaction. Spectral data follow. ¹ H NMR (CDCl₃) δ 3.3 (triplet, 2H),1.55 (t, 2H), 1.35 (bs, >20H).

EXAMPLE 15 4-Hydroxy-2-oxo-2.5-dihydro-furan-3-carboxylicacid-2-cyclododecyl-ethylamide

To a dry flask containing 40 mL of chloroform was added 1.87 g (18.7mmol) of tetronic acid, 2.29 g (18.7 mmol) of4-N,N-dimethylaminopyridine, 2.6 mL (18.7 mmol) of triethylaminefollowed by 4.4 g (18.7 mmol) of 2-cyclododecylethyl isocyanate. Thereaction mixture was allowed to stir overnight at room temperature,after which time a small amount of starting materials was still evidentby TLC. The reaction mixture was quenched with 1N HCl, the layersseparated, the organic fraction dried over MgSO₄, filtered, and strippedof solvent. The residue was flash chromatographed using acidic silicagel eluting with 25% EtOAc/hexane to give a yellow solid. M.P. 135°-137°C. Spectral data follow: ¹ H NMR (DMSO-d₆) δ 8.05 (bs, 1H), 4.6 (s, 2H),4.2 (bs, 10H), 3.25 (t, 2H), 2.5 (s, 3H), 1.2-1.4 (bm, >20H); IR (KBr)3340, 2920, 2850 (C--H), 1750, 1650 cm⁻¹ (C=0);

    ______________________________________                                                         Theory                                                                              Found                                                  ______________________________________                                        Analysis: % C          67.63   66.78                                                    % H          9.26    8.89                                                     % N          4.15    3.97                                           ______________________________________                                    

EXAMPLE 16A

Cyclododecylmethyl 3 allyl ether

To 2.4 g (50.4 mmol) of 50% sodium hydride that had been washed twicewith hexanes was added 150 mL of N,N-dimethyl formamide followed by 10.0g (50.4 mmol) of cyclododecyl methanol. The reaction mixture was stirredat room temperature for several hours and then at 65° C. until theevolution of gas had ceased. 8.7 mL (100.8 mmol) of allyl bromide wasthen added and the reaction mixture was allowed to stir overnight at 65°C. The reaction was quenched with water, extracted 3 times with ether,the combined ether layers was washed with water, dried over MgSO₄ andevaporated. The residue was flash chromatographed on silica gel using 2%Et₂ O/Hexanes to give 5.5 g of an oil (49%). NMR(CDCl₃): 5.9 ppm (m,1H); 5.2 (m, 2H); 3.95 (d, 2H, J=7 Hz); 3.25 (d, 2h, J=8 Hz); 1.3 (m,>10H).

EXAMPLE 16B Cyclododecylmethoxy acetic acid

A solution of 4.5 g (20.1 mmol) of cyclododecylmethyl allyl ether inacetone at -78° C. was subjected to ozonolysis until the solutionremained blue. The reaction mixture was quenched by the addition of 3.0mL (40.2 mmol) of dimethyl sulfide. The volatiles were evaporated andthe residue was subjected to Jones oxidation in acetone at)°C. Thereaction mixture was partitioned between ether and 1N HCl the aqueouslayer was washed with ether and the combined ether layers were driedover MgSO₄ and evaporated. The residual material was recrystallized fromhexanes to give 2.0 g of white solid (40%). NMR (CDCl₃): 4.0 ppm (s,2H); 3.4 (d, 2H, J=7 Hz); 1.4 (m, >10H).

EXAMPLE 16C

According to the methods and reagents described above,3-[(cyclododecylmethoxy)acetyl]-4-hydroxy-2(5H)-thiophenone was preparedas an off-white solid having a melting point of 93°-94° C.

EXAMPLE 17

The compounds of the invention are tested in an in vitro phospholipaseA₂ assay to determine the ability of the compounds to inhibit thebiosynthesis of platelet-activating factor and LTB₄ in purified humanneutrophils.

This assay is carried out as follows:

Isolation of Human Polymorphonuclear Neutrophils

A leukocyte enriched blood sample obtained from a healthy male donor isprocured by leukophoresis using a Haemonetics model 30+ blood processor(Biological Specialties, Inc., Lansdale, Pa.). The top "platelet-rich"layer is removed after a low speed spin (35×g, 15 min, 25° C.) of thesample. The remaining cell suspension is centrifuged (400×g, 10 min, 25°C.) to sediment the remaining cells. The supernatant is discarded andthe cell pellet resuspended in 120 ml HBSS (without Ca⁺⁺ /Mg⁺⁺). Thecell suspension is subjected to ficoll-hypaque sedimentation (Histopaque1077,400×g, 30 rain, 25° C.). Contaminating erythrocytes are lysed byhypotonic shock (1 min). The cells are then washed once with 40 ml ofHBSS and resuspended with HBSS (without Ca⁺⁺ /Mg⁺⁺) to a concentrationof 2.5×10⁷ cells/ml for further use. Greater than 95% purity isobtained, as assessed by microscopic examination.

Platelet-Activating Factor Biosynthesis in Human PolymorphonuclearNeutrophils (PMN)

One ml of human PMN (2.5×10⁷ cells/ml) is incubated with vehicle ordrugs (10 μl) for 10 minutes at 30° C. After preincubation, an equalvolume of HBSS (1 ml) containing 2.4 mM CaCl₂, 6 gM calcium ionophoreA23187 and 50 μCi [³ H]-acetate is then incubated at 30° C. for 15minutes. An aliquot (100 μl) of the reaction mixture is taken out andmixed with 900 μl of 15% ethanol. LTB₄ is extracted by using solid phaseextraction on reverse phase C18 columns to remove excess [³ H]-acetateand PAD. The C₁₈ column is prewashed once with 2 ml of ethanol andwater. The sample aliquot is acidified with 0. 1N HCl to pH 3 beforeapplying to the column. The column is then washed with 2 ml of waterfollowed by 2 ml of 15% ethanol and 2 ml of petroleum ether to removeexcess labeled acetate. The sample is eluted with 2 ml of ethyl acetate.The collected samples are dried with nitrogen and resuspended in 0.5 mlRIA buffer. The quantity of LTB₄ in the sample is obtained from RIAdetermination. For PAF determination, the reaction is terminated byaddition of 5 ml chloroform:methanol:acetic acid (2:1:0.04, v/v/v). [³H]-PAF is obtained by Bligh and Dyer extraction. The chloroform phase isremoved and dried under nitrogen. The residue is redissolved in 75 μl ofchloroform:methanol (80:20, v/v). [³ H]-PAF is resolved from otherphospholipids by TLC on RPC₁₈ plates with a solvent system ofchloroform:methanol:water (2:3: 1, v/v/v) and is quantitated using aBerthold automated TLC linear analyzer.

Data presented are the mean+/-s.d. of the values relative to controlA23187 stimulated cells for each experiment assayed in triplicate.Percent inhibition when used is calculated as:

    % Inhibition=100-[(x÷Control)×100]

Dose response analysis is performed by non-linear regression analysisfor curve fitting and IC₅₀ determination.

In this assay, scalaradial, an irreversible inhibitor of PLA₂, isolatedfrom the marine sponge Cacospongia sp. gives an IC₅₀ of 1.0 μM.

When tested in this assay, the compounds of the invention gave thefollowing results:

                  TABLE I                                                         ______________________________________                                        Compound of                                                                            Dose,    % Inhibition                                                                             Dose,  % Inhibition                              Example No.                                                                            μM    PAF        μM  LTB.sub.4                                 ______________________________________                                        1        10       84.3       10     95.1                                      2        10       79.2       10     69.6                                      3        10       64.4       10     90.4                                      ______________________________________                                    

EXAMPLE 18

The compounds of the invention are tested in an in vitro isolatedphospholipase A₂ assay to determine the ability of the test compounds toinhibit the release of arachidonic acid from an arachidonicacid-containing substrate by the action of phospholipase A₂ enzyme fromhuman and non-human sources.

This assay is carded out as follows:

Into a 15 mL polypropylene tube are added the following:

    ______________________________________                                        Agent          Volume, μL                                                                             Final Conc.                                        ______________________________________                                        .sup.3 H-AA E. coli substrate.sup.1                                                          25          5 nmoles PL                                        CaCl.sub.2 (0.1M).sup.2                                                                       5          5 mM                                               Tris-HCl (0.5M) pH 7.5.sup.3                                                                 20          100 mM                                             Water.sup.4    25                                                             Drug/vehicle.sup.5                                                                            1          50 μM                                           PLA.sub.2.sup.6                                                                              25          Volume yielding 12%                                                           hydrolysis in 10 min.                                             100                                                            ______________________________________                                         *preincubate at room temperature 30 min prior to substrate addition.          .sup.1 Prepared by adding 2 mL deionized and distilled water to 2 mL          .sup.3 Harachidonate labeled E. coli (lower count), to which is added 1 m     of .sup.3 Harachidonate labeled E. coli (higher count) to yield a total o     5 m substrate (containing 1000 nmoles phospholipid).                          .sup.2 Stock 0.1 m CaCl.sub.2, required for enzyme activity.                  .sup.3 Stock 0.5 m TrismaBase.                                                Stock 0.5 M TrismaHCl. Adjust pH to 7.5 (optimum for enzyme).                 .sup.4 Deionized and distilled water.                                         .sup.5 Stock 10 mM prepared in dimethyl sulfoxide. Make 1:2 dilution with     dimethyl sulfoxide and add 1 μL to 100 μ L assay tube.                  .sup.6 Two human PLA.sub.2 enzymes are used:                                  a) Semipurified human platelet acid extract PLA.sub.2 (in 10 mM sodium        acetate buffer, pH 4.5). Remove protein precipitate by centrifugation at      about 2200 rpm for 10 minutes.                                                b) Purified human synovial fluid.                                        

Incubate the 100 μL reaction mixture for 10 minutes at 37° C. in ashaking water bath. The reaction is terminated by the addition of 2 m.Ltetrahydrofuran, followed by vortexing. NH₂ columns (100μg/mL--Analytichem International) are conditioned with 0.5 mLtetrahydrofuran followed by 0.5 mL tetrahydrofuran/water (2 mL:0.1 mL,v/v).

The sample is loaded onto the columns and slowly drawn through them. Thehydrolyzed arachidonic acid retained in the columns is eluted therefromwith 1 mL tetrahydrofuran/glacial acetic acid (2%). The arachidonic acidis transferred to scintillation vials and quantitated by β-countinganalysis. A "total counts" sample is prepared by pipetting 25 μL ³H-arachidonate E. coli directly into a scintillation vial to which isadded 1 mL tetrahydrofuran. 10 mL aquasol (scintillation cocktail) isadded to all samples. ##EQU1##

    ______________________________________                                        Activity of Standard Drugs:                                                               IC.sub.50 (μM)                                                               Human Platelet                                                                            Human Synovial                                      Drug          PLA.sub.2   PLA.sub.2                                           ______________________________________                                        Arachidonic Acid                                                                            8.6         3.2                                                 Monoslide     25.2        0.14                                                ______________________________________                                    

When tested in this assay, the compounds of the invention gave thefollowing results:

                  TABLE II                                                        ______________________________________                                        Compound of      % Inhibition at 10 μM                                     Example No.      HP*     HSF**                                                ______________________________________                                        1                -3.7    41.9                                                 2                18.2    53                                                   ______________________________________                                         *human platelet                                                               **human synovial fluid                                                   

EXAMPLE 19

The ability of the compounds of the invention to act as inhibitors ofthe enzymes 5-lipoxygenase and cyclooxygenase is measured in theresident murine peritoneal macrophage assay.

This assay is carried out as follows:

Resident peritoneal macrophages are collected from female Swiss Webstermice (49 days old, 20-25 gins, Buckshire) by lavaging with 7-8 ml HanksBalanced Salt Solution (HBSS) without Ca⁺⁺ and Mg⁺⁺ (GIBCO). The lavagefluid from several mice is pooled and centrifuged at 4° C. for 10minutes at 400×g. The cell pellet is resuspended in Medium 199 (GIBCO)with HEPES buffer containing 100 μg/ml gentamicin. Two ml of the cellsuspension (4×10⁶ cells) are then plated on 35 mm culture dishes (Nunc).

A macrophage monolayer is established after a 1-1.5 hour incubation ofthe cells at 37° C. in an atmosphere of 95% O₂ and 5% CO₂. Themonolayers are washed 2× with 2 ml HBDSS, containing Ca⁺⁺ and Mg⁺⁺ afterwhich 2 ml Medium 199 supplemented with 10% freshly thawedheat-inactivated fetal bovine serum and 100 μg/ml gentamicin is addedfor an overnight incubation.

Residual serum and cellular debris are removed from the monolayers bywashing 3× with 2 ml HBSS containing Ca⁺⁺ and Mg⁺⁺. Macrophages arepreincubated for 5 minutes with 1 ml serum-free M199 containing 10 μldimethyl sulfoxide (DMSO) vehicle or test compound prior to cellactivation with zymosan (100 Mg/ml) or arachidonic acid (AA) (2 μM).After 2 hours, the supernatants are removed and either assayed for LTC₄and PGE₂ by radioimmunoassay (RIA) directly or stored at -20° C. In allcases, results are expressed as ng metabolite/4×10⁶ cells.

Summary of RIAs used for quantitation of metabolite levels in zymosan orarachidonic acid stimulated mouse macrophage culture media.

    ______________________________________                                                                 Metabolite Levels                                                Range of     (ng/4 × 10.sup.6 cells)                        Metabolite  detection (μg/ml)                                                                       (x ± S.E.M..n)                                    ______________________________________                                        LTC.sub.4    0.25-16     93.7 ± 9.9 (34)                                   PGE.sub.2   0.027-20     30.90 ± 1.93 (39)                                 ______________________________________                                    

Calculations

Raw data (dpm) may be stored directly onto an "Autostart" tape using theHP85 in room C-096. The raw data are converted to ng metabolite/4-10⁶cells using the standard curve by a "RIANAL" program (HP85) or a"NONLIN" program (HP9816). Results are then expressed as percentinhibition of zymosan induced, leukotriene or prostaglandin synthesis(control) using the equation: ##EQU2##

REFERENCE COMPOUNDS

The compounds used are listed below. IC₅₀ values of reference5-lipoxygenase and/or cyclooxygenase inhibitors.

    ______________________________________                                        IC.sub.50 μM (95%) Confidence limits)                                      Compound      LTC.sub.4     PGE.sub.2                                         ______________________________________                                        BW 755c       0.21          1.04                                                            (0.10, 0.42)  (0.73, 1.49)                                      ETYA          0.44          1.26                                                            (0.36, 0.53)  (0.99, 1.60)                                      Indomethacin  >50           0.002                                                                         (0.001, 0.003)                                    NDGA          1.87          2.15                                                            (0.22, 15.57) (1.15, 4.04)                                      ______________________________________                                    

When tested in this assay, the compounds of the invention exhibited thefollowing levels of enzyme inhibition:

                  TABLE III                                                       ______________________________________                                        Compound of                                                                            LTC.sub.4       PGE.sub.2                                            Example No.                                                                            Dose μM                                                                             % Inhibition                                                                             Dose μM                                                                           % Inhibition                              ______________________________________                                        1        0.5      29.0       0.5    46.7                                      2        0.5      76.1       0.5    97.1                                      3        0.5      66.2       0.5    75.9                                      15       0.5      25.9       0.5    52.0                                      16       0.5      33.0       0.5    77.0                                      ______________________________________                                    

What is claimed is:
 1. A compound having the formula ##STR13## wherein X is NR;R is hydrogen or lower alkyl; R¹ and R² are each, independently, hydrogen, C₁ -C₁₀ alkyl, C₃ -C₂₀ cycloalkyl, phenylloweralkyl, or substituted phenylloweralkyl substituted by halo, lower alkyl, lower alkoxy, halo lower alkyl, amino, monoloweralkylamino, diloweralkylamino or sulfonamido; A is O, S ,NR, or a chemical bond; m is0-15; n is 0-20; p is0-15, where m+p≦15;and the pharmacologically acceptable salts thereof.
 2. The compound of claim 1, having the name 3-(3-cyclododecyl-1-oxopropyl)-1,5-dihydro-4-hydroxy-1-methyl-1H-pyrrolin-2-one.
 3. A method for treating immunoinflammatory conditions in mammals which comprises administering to a mammal so afflicted an effective amount of a compound having the formula ##STR14## wherein X is NR;R is hydrogen or lower alkyl; R¹ and R² are each, independently, hydrogen, C₁ -C₁₀ alkyl, C₃ -C₂₀ cycloalkyl, phenylloweralkyl, or substituted phenylloweralkyl substituted by halo, lower alkyl, lower alkoxy, halo lower alkyl, amino, monoloweralkylamino, diloweralkylamino or sulfonamido; A is O, S ,NR or a chemical bond; m is 0-15; n is 0-20; p is0-15, where m+p≦15;and the pharmacologically acceptable salts thereof. 